Project Summary The proper distribution of retinal ganglion cell (RGC) axons at the optic chiasm to the same (ipsilateral) and opposite (contralateral) sides of the brain is crucial for proper binocular vision. Our work has defined axon guidance receptors and transcription factors directing the ipsilateral and contralateral pathways in pigmented animals. We have shown that a molecular program of transcription and guidance factors, including Foxd1, Zic2, and EphB1, is essential for directing the ipsilateral retinal projection, and a complex of factors including guidance molecules Nr-CAM, Plexin-A1, and Semaphorin6D regulate the contralateral projection. These genes control RGC axon growth from the eye as RGC axons cross or avoid the chiasm midline, and subsequently innervate the thalamus and superior colliculus. Albinism is characterized by perturbed melanogenesis in the skin and the eye and results in fewer RGCs that project ipsilaterally and, consequently, impaired stereovision. The molecular mechanisms that link altered melanin biogenesis in the RPE with the reduction in ipsilaterally-projecting (i) RGCs are not understood. However, our studies have indicated that in the albino, neurogenesis in the ventrotemporal (VT) retina, where iRGCs originate, is temporally shifted, and that fewer RGCs express genes such as Zic2 that specify an ipsilateral trajectory. These findings provide new inroads on how to approach the albino mystery. We hypothesize that factors related to melanin production in the normally pigmented RPE influence the development of RGCs in the neural retina, namely their genesis and fate specification. Based on our new RPE studies and those of other labs, it is likely that such factors could affect RGC development via junctions at the RPE-retina interface, where we have identified alterations in the albino. In addition, some RPE factors, missing or upregulated in the albino, are likely to be important for RPE cell health. In the proposed studies, Aim 1 will identify genes that are differentially expressed in pigmented and albino RPE during the period of iRGC genesis, by conducting candidate-based and unbiased gene profiling analyses, followed by validation of these genes by immunostaining, in situ hybridization and qPCR. In Aim 2, we will test a role for select factors on RGC subtype specification through in vitro assays of RGCs alone, RPE alone, and in organotypic slice cultures. By addition of proteins or exogenous gene expression, we will determine if candidate RPE factors can rescue or induce albino retinal and RPE phenotypes. Identification of RPE-derived factors that influence RGC subtype specification will reveal ocular tissue interactions during vision system development, that relate to human disease. These studies will also illuminate how neuronal differentiation programs govern axon projection and connectivity in the nervous system.